This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. Notch proteins are the receptors in a highly conserved signal transduction system used to communicate signals between cells that contact each other. These signals influence a wide spectrum of cell fate decisions, both during development and in the adult organism. However, dysregulated signaling has also been implicated in a number of different human diseases ranging from neurodegeneration to cancer. The goal of these studies is to use X-ray crystallography to help elucidate the molecular basis for Notch signal transduction. Notch receptors are single-pass transmembrane proteins normally activated by ligand-induced proteolysis. Prior to ligand binding, Notch receptors are normally maintained in a quiescent and proteolytic resistant state by a negative regulatory region (NRR), or "activation switch," which includes three LNR repeats and the heterodimerization domain divided by cleavage at S1 during maturation see Figure 1). Though Notch proteins reach the cell membrane in a conformation resistant to proteolytic activation, they become sensitive to ADAM metalloproteases such as Kuzbanian (ADAM10) and TNFa converting enzyme (TACE or ADAM17) upon binding a ligand of the Delta, Serrate, or Lag-2 (DSL) family. One major area of interest has been to elucidate the structural basis for Notch autoinhibition, and determine the nature of the conformational change that enables proteolysis in response to ligand stimulation. After metalloprotease cleavage, the truncated transmembrane fragment of Notch undergoes further proteolysis by the multiprotein gamma-secretase complex, which releases the intracellular portion of Notch (ICN) from the membrane. ICN then enters the cell nucleus, where it assembles a multiprotein transcriptional activation complex that results in the transcription of notch-responsive genes. A second focus of our research is to determine the structural basis for the formation of Notch transcription complexes, and for the cooperative assembly of higher order complexes on target DNA.